We propose to use a genetic approach to investigate the influence of differences in enzymes of the testosterone biosynthetic pathway on the capacity of Leydig cells to produce testosterone. To accomplish this objective a variety of experiments have been designed using purified Leydig cells from numerous strains of inbred mice (progenitor strains), as well as various recombinant inbred (RI) lines derived from these progenitor strains. We will survey first for inbred strains which differ significantly in Leydig cell capacity for testosterone production. We will also survey for differences in the level of activities of the testosterone biosynthetic enzymes associated with the smooth endoplasmic reticulum (SER), 3Beta-hydroxysteroid dehydrogenase-isomerase, 17Alpha-hydroxylase, C17-20 lyase and 17-ketosteroid reductase. The appropriate RI lines of the progenitor strains with the greatest interstrain differences for maximum testosterone production and maximal activity of one or more of the enzymes will be analyzed for cosegregation of phenotypes. The RI lines will also be used to estimate the number of genes responsible for maximum testosterone production, the chromosomal location of the genes encoding the enzymes and possible linkage of the genes encoding the enzymes. Finally, we will survey numerous inbred strains of mice for enzymes which differ with regard to clearly detectable qualitative characteristics such as thermostability or electrophoretic mobility. Such differences in physical characteristics are indicative of structural gene variants. This approach should establish, without resorting to environmental perturbation, whether genetically determined variations in the level of activity of any one of the enzymes of the enzymes of the SER result in a change in the capacity of Leydig cells to produce testosterone.